Project description:The goal of this study is to compare the transcriptome stability of DN1wt versus DN1gl/gl and DN1CD2-Ostm1gl/glTR Methods: DN1 thymocytes mRNA profiles of 19-day-old wild-type (WT), osteopetrotic grey-lethal (gl/gl) and transgenic CD2-Ostm1 gl/gl mice were generated by deep sequencing,DN1 cells from 2-3 mice per genotype were pooled, no technical replicates, using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: TopHat and Cufflinks. qRTâ??PCR validation was performed using SYBR Green assays Results: DN1 gl/gl showed a distinctive transcriptome signature in comparison to DN1 wt and DN1 gl/glTR with respectivelly 146 and 205 differentially expressed genes and 205. Migration genes were significantly affected in DN1 gl/gl samples and RAC1 and S1PR1 gene expressions were confirmed using RT-qPCR. Conclusions: RNA seq allowed us to identify the enhanced expression of migration genes (RAC1 and S1PR1) in DN1gl/gl, that were both normalized in the DN1 cells from transgenic gl/glTR mice, which suggests defective T cell migration associated to the osteopetrotic thymus phenotype. DN1 thymocytes mRNA profiles of 19-day-old wild-type (WT), osteopetrotic grey-lethal (gl/gl) and transgenic CD2-Ostm1 gl/gl mice were compared by deep sequencing,DN1 cells from 2-3 mice per genotype were pooled, no technical replicates, using Illumina HiSeq 2000.
Project description:The goal of this study is to compare the transcriptome stability of DN1wt versus DN1gl/gl and DN1CD2-Ostm1gl/glTR Methods: DN1 thymocytes mRNA profiles of 19-day-old wild-type (WT), osteopetrotic grey-lethal (gl/gl) and transgenic CD2-Ostm1 gl/gl mice were generated by deep sequencing,DN1 cells from 2-3 mice per genotype were pooled, no technical replicates, using Illumina HiSeq 2000. The sequence reads that passed quality filters were analyzed at the transcript isoform level with two methods: TopHat and Cufflinks. qRT–PCR validation was performed using SYBR Green assays Results: DN1 gl/gl showed a distinctive transcriptome signature in comparison to DN1 wt and DN1 gl/glTR with respectivelly 146 and 205 differentially expressed genes and 205. Migration genes were significantly affected in DN1 gl/gl samples and RAC1 and S1PR1 gene expressions were confirmed using RT-qPCR. Conclusions: RNA seq allowed us to identify the enhanced expression of migration genes (RAC1 and S1PR1) in DN1gl/gl, that were both normalized in the DN1 cells from transgenic gl/glTR mice, which suggests defective T cell migration associated to the osteopetrotic thymus phenotype.
Project description:In this study, to investigate the pathogenic role of transcriptional regulator LMO2 during T lineage development, we isolated DN1, DN3, DP, CD4SP, CD8SP thymocytes, splenic CD4+ T cells and splenic CD8+ T cells from wild type and LMO2 over-expressing C57BL/6J mice for RNA-seq, and DN3 (CD25+), DP thymocytes, splenic CD4+/CD8+ T cells from transgenic mice and wild type DN3 (CD25+) thymocytes for ChIP-seq.
Project description:Mutations in the isocitrate dehydrogenase 1 (IDH1) and IDH2 genes are frequently observed in a wide variety of hematologic malignancies, including myeloid and T-cell leukemias. In this study, we generated Idh2R140Q transgenic mice to examine the role of the Idh2R140Q mutation in leukemia. No leukemia developed in Idh2R140Q transgenic mice, suggesting a need for additional genetic events for leukemia development. Since myeloid cells from NUP98-HOXD13 fusion (NHD13) transgenic mice frequently acquire somatic Idh mutations when they transform to AML, we generated Idh2R140Q/NHD13 double transgenic mice. Idh2R140Q/NHD13 transgenic mice developed an immature T cell leukemia with an immunophenotype similar to double-negative 1 (DN1) or DN2 thymocytes. Idh2R140Q/NHD13 leukemic cells were enriched for an early thymic precursor transcriptional signature, and the gene expression profile for Idh2R140Q/NHD13 DN1/DN2 T-ALL closely matched that of human early/immature T cell precursor (EITP) ALL. Moreover, recurrent mutations found in EITP ALL patients, including KRAS, PTPN11, JAK3, SH2B3, and EZH2 were also found in Idh2R140Q/NHD13 DN1/DN2 T-ALL. In vitro treatment of Idh2R140Q/NHD13 thymocytes with enasidenib, a selective inhibitor of mutant IDH2, led to a marked decrease in leukemic cell proliferation. These findings demonstrate that Idh2R140Q/NHD13 mice can serve as a useful in vivo model for the study of EITP ALL development and therapy. We used gene expression arrays to compare the global gene expression profiles between Idh2R140Q/NHD13 DN1/DN2 T-ALL, non-DN1/DN2 cortical T-ALL (CD4+CD8+) and wild-type thymus.
Project description:Identify the function of pE66L Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and Nrl-- Retinal Transcriptomes
